S 4700 cold cathode field emission sem

All particle characterization methods were completed as previously reported [22 (link), 27 (link), 35 (link)]. Briefly, all particles were deemed to have undetectable endotoxin (<0.125 EU/mg) per the LAL Chromogenic Endotoxin Quantitation Kit (Waltham, MA). For imaging by scanning electron microscopy (SEM), particles were cast on an aluminum stub (Ted Pella, Redding, CA), and after being sputter coated with palladium, were imaged on a S-4700 Cold Cathode Field Emission SEM (Hitachi, Tokyo, Japan). The particle hydrodynamic size by volume was acquired using a Nanobrook 90Plus Zeta (Brookhaven Instruments Corporation, Holtsville, NY). cGAMP loading was determined as previously described[22 (link)] using a high-performance liquid chromatography (HPLC) method by detecting cGAMP at 254 nm. Briefly, HPLC was run using an isocratic mobile phase (80% water:20% methanol v/v) at a rate of 0.6 mL/min through a Aquasil C18 (Thermo Fisher Scientific, Waltham, MA). To determine the cGAMP release profile, the particles were incubated in PBS in the presence or absence of 10% v/v FBS pH 7.4 at 37 °C and at predetermined time points the same HPLC method was used to measure percent cGAMP released.

Cross-sectioned HCCS-PDA scaffolds were fixed on a holder, sputter-coated with platinum (4 nm) in a vacuum and imaged using a Hitachi S-4700 Cold Cathode Field Emission SEM (Hitachi High Technologies America, Inc., Schaumburg, IL USA). The scaffolds were fixed in 10% neutral formalin for 24 hours, washed 3 times with deionized water, and followed by critical point drying process using EMS 850 Critical Point Dryer (Electron Microscopy Sciences, Hatfield, PA USA) before acquiring image using the SEM.